KR101398591B1 - Camptothecin derivatives with antitumor activity - Google Patents

Abstract

Novel camptothecin derivatives having antitumor activity, the processes for the preparation thereof, the use thereof as antitumor drugs and pharmaceutical compositions containing them.

Description

Camptothecin derivatives with antitumor activity < RTI ID = 0.0 >

The present invention relates to novel camptothecin derivatives with anticancer activity, processes for their preparation, their use as anti-cancer drugs and medicinal compositions comprising them.

Camtothecine was first described as Camptotheca acuminata (N apple, Nyssaceae) as described in "Wall and Wani" (1966, 88, 3888-3890) ≪ / RTI > Camptothecin has a broad spectrum of cytotoxic activity, particularly in colorectal tumors and other solid tumors and leukemia, but is not used in therapy due to its high toxicity. This problem is particularly prevalent in hemorrhagic cystitis, stomach toxicity and myelosuppression.

Many camptothecin analogs have been synthesized to obtain compounds with low toxicity and high solubility. Currently, two drugs called so-called CPT-11 and topotecan are used in clinical practice. But are not limited to, belotecan, rubitecan, exatecan, gimatecan, pegamotecan, lurtotecan, karenitecin, Other derivatives such as afeletecan, homocamptothecin, diflomotecan and many other substances have undergone clinical trials. Compound CPT-11 is approved for the treatment of many solid tumors and ascites (rectal, skin, gastrointestinal, lung, cervical, ovarian, non-Hodgkin's lymphoma) Is a prodrug with high solubility for the 10-hydroxy-7-ethyl camptothecin (commonly known as SN-38).

Topotecan can be used to treat tumors of the lungs, stomach, liver, ovaries, chest, prostate, esophagus, rectum, soft tissues sarcomas, head and neck, glioblastoma, chronic and acute myelocytic leukemia And dissolves in a physiological solution having an activity against water. However, topotecan causes serious side effects such as neutropenia and thrombocytopenia.

Rutotecan is a more soluble derivative that is active in tumors of the neck, ovaries, chest, rectum, and lung microcytoma. However, rutotecan also has blood toxicity.

Rubitecan is a prodrug for effective oral use against tumors of the pancreas, ovary and chest.

Camptothecin and its analogs are effective against tumors resistant to conventional drugs, including topoisomerase II inhibitors, as do all topoisomerase I inhibitors; Maintains high levels of topoisomerase during the entire cell cycle; Do not induce multiple-drug resistance (Pgo or MRP) or detoxification metabolism mediated by the enzyme.

Recent studies have focused on novel inhibitors of topoisomerase I that have lower toxicity than the currently used drugs.

Open-ring camptothecin derivatives exhibit high protein binding (especially with albumin) and low distribution in tumor tissues. As a result, the product accumulates in the body and the tumor is badly affected.

In contrast, the high lipophilicity of the lactone form promotes adhesion of camptothecin derivatives to cell membranes, particularly red blood cells, and affects the tissue / plasma distribution ratio. For this reason, the research has focused on two alternative approaches: a) the design of low protein binding products still having good solubility; b) Very well designed products with therapeutic effects even at extremely low doses.

The modifications at the 7-, 9-, 10- and 11-positions have generally been shown to be well tolerated without affecting the stability of the complex of the DNA-topoisomerase I-camptothecin triple atom . The formation of the complex is responsible for the anti-cancer activity of the compounds.

Products with a 20R configuration have proved to be inactive or have very low activity compared to products with a 20S configuration. It is also consistent with its natural composition.

In general, modification at the 5-position is considered undesirable for the formation of a triple atom complex. On the other hand, modifications in the pyridone rings D and E have been reported to be detrimental to the activity of the product.

In a first aspect, the invention relates to a camptothecin derivative of general formula I:

(I)

Wherein R is alkyl, aminoalkyl, hydroxyalkyl, nitrile, alkoxymino, aryloxymino, silylalkyl;

R1 is hydrogen, hydroxy, alkoxy, aminoalkyl;

R2 is hydrogen, hydroxy, alkoxy, aminoalkyl, or optionally protected hydroxyl;

Wherein the alkyl, alkoxy, aminoalkyl or alkoximino groups may contain from 1 to 8, preferably from 1 to 4, carbon atoms in the form of a straight or branched chain. On the other hand, the aryloximino group may contain 5 to 10 carbon atoms;

Also encompassed by the present invention are the above pharmaceutically acceptable salts, isomers, enantiomers, diastereomers and corresponding mixtures of the compounds of formula (I).

The compounds of the invention show high efficacy even at low protein binding, good solubility and very low usage.

The preferred synthetic routes for the preparation of the compounds of the invention are described in the following tables, which substantially comprise the following steps:

a) protection of the precursor hydroxy groups;

b) derivatization at the 5-position with N, N-diprotected hydrazine;

c) selective conversion of a pyridone ring to a thiopyridone ring;

d) elimination of the protecting groups by concomitant cyclization;

e) selective aromatization of the pyrazole ring;

In the above table, R, R 1 and R 2 have the meanings as described above, and PG is a hydroxy-protecting group.

The hydroxyl is protected by an easily cleavable acyl group, preferably trichloroacetate and Troc or a silyl group, preferably triethylsilyl.

Induction at the 5-position using protected hydrazine can be accomplished by treating the precursor with a strong organic base such as LiHMDS and converting the resulting carbanion into di-t-butoxy With an aza dicarboxylate such as an azodicarboxylate or azodicarboxylate or dibenzyloxy azadicarboxylate. The term " anhydride " The conversion of the pyridone ring to the thiopyridone ring is achieved by reacting 2,4-bis (4-methoxyphenyl) -1,2,3,4-dithiaphosphethane-2,4-disulfide 1982, 1, 69; Yde B (1982), which is known as Lawsson's remedy (Cava PM et al., Tetrahedron 1985, 41, 5061; Cherkasov RA et al Tetrahedron 1985 41, 2567; Ghattas AAG et al, Sulfur Lett. et al., Tetrahedron 1984, 40. 2047) or an equivalent reagent. Lawson's reagent is preferred.

The purpose of selective conversion to thiopyridone is to promote ring closure once the hydrazine is unprotected. However, it has been observed that the closure reaction occurs spontaneously and immediately without the activation of pyridinecarbonyl, for example with thiocarbonyl.

When the hydroxy-protecting groups are silyl and those at the nitrogen position are carbamates, they are usually removed by trifluoroacetic acid. In an alternative process, steps b) and c) may be reversed.

The compounds of the present invention are tested in cytotoxicity analysis on a broad spectrum of tumor cells. For example, cytotoxicity on the NCI-H460 cell line (NSCL cancer) associated with two compounds of formula (I) has been reported to use drugs such as camptothecin and topotecan and SN-38 as standard:

The most active compounds are evaluated by DNA cleavage analysis, which measures active condensation and impairment of persistence (see the Examples section). The derivatives of formula (I) surprisingly show a higher persistence in inhibiting DNA replication while maintaining effective cytotoxic activity than reference standards (particularly topotecan and camptothecin).

In another aspect, the invention relates to medicinal compositions comprising a compound of formula (I), together with pharmaceutically acceptable carriers and excipients. The forms of the medicament suitable for oral or parenteral administration of the compounds (I) are preferably solid, such as capsules, tablets and granules, or preferably injectable or infusible, Lt; / RTI > solution.

The suitably formed compounds of the present invention are useful for the treatment and / or prophylaxis of solid tumor and leukemia, especially lung, ovarian, breast, gastric, liver, prostate, soft tissue sarcoma, head and neck, esophagus, pancreas, colon, rectum, glioblastoma, , ≪ / RTI > and the like.

Examples

Example I: 20-OTES-camptothecin

Camptothecin (0.100 g, 0.287 mmol) was suspended in anhydrous dimethylformamide (3 mL) under an inert atmosphere and the resulting suspension was loaded with imidazole (0.980 g, 1.44 mmol) is added. The mixture was stirred for 10 minutes and then triethylsilyl chloride (TES-Cl) (0.193 mL, 1,15 mmol) was added thereto followed by 4-dimethylaminopyridine (DMAP) g 0.287 mmol) is added. After 46 hours, the reaction mixture is evaporated under vacuum (TLC control, complete elution of the reagent, eluent CH ₂ Cl ₂ / MeOH = 30/1). The solid is then redissolved in CH ₂ Cl ₂ and washed with H ₂ O and saturated NH ₄ Cl. The liquid phase is extracted using CH ₂ Cl ₂ (2 × 10 mL). The organic phases are combined on Na ₂ SO ₄ and dried, filtered under vacuum and condensed. Thus, the desired product (0.133 g, 0.287 mmol) was obtained as a light yellow solid.

_{^{1 H NMR (CDCl 3, 400}} MHz) δ 8.37 (s, 1 H, Ar, H-7), 8.25 (d, 1 H, J = 8.4 Hz, Ar), 7.92 (d, 1 H, J = 8.0 Hz), 7.82 (t, 1H, J = 8.0 Hz, Ar), 7.65 (t, 1H, J = 8.4 Hz, Ar), 7.57 , 1 H, J = 16.4 Hz , H-17), 5.29 (s, 2 H, H-5), 5.25 (d, 1 H, J = 16.4 Hz, H-17), 2.00-1.84 (m, 2 H, H-19), 1.03-0.93 (m, 12H), 0.80-0.71 (m, 6H). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 171.7, 157.6, 152.5, 151.5, 149.0. 145.9, 130.9, 130.4, 130.0. 128.4, 128.1, 128.0. 127.9, 118.9, 94.4, 75.3, 66.0. 50.0. 33.2, 7.9, 7.2, 6.4.

Example II

: Preparation of 5-di-t-butoxycarbonylhydrazino-20-OTES-camptothecin

The camptothecin 20-OTES (0.100 g, 0.216 mmol) was dissolved in anhydrous THF (6 mL) with stirring under an inert atmosphere and then cooled to -78 ° C and 1.0 M LiHMDS in THF (0.281 mL, 0.281 mmol) The solution is put into it. After 20 minutes, di-tert-butylazo dicarboxylate (DTBAC) (0.075 g, 0.324 mmol) in anhydrous THF (2 mL) is added. After 4 h at -78 [deg.] C, the disappearance of the reagent is monitored by TLC (hexane / AcOEt = 3/1). The formation of the two diastereomers is observed. The reaction is cooled by the addition of saturated NH ₄ Cl. The liquid phase is extracted using CH ₂ Cl ₂ (3 x 15 mL) and the organic phases are combined, dried over Na ₂ SO ₄ , filtered under vacuum and condensed. The residue is purified by flash chromatography (SiO _2, hexane / AcOEt = 3/1), thus to obtain a mixture of the two the two isomers (0.145 g, 0.210 mmol, 97 %). The two isomers are separated by further chromatography.

Sequence of elution:

The first _{^{diastereomer: 1 H NMR (CDCl 3,}} 400 MHz) δ 8.80 (br s, 1 H, Ar), 8.23 (d, 1 H, J = 8.4 Hz, Ar), 8.01 (br d, 1 H, H-5), 5.61 (d, 1H, Ar), 7.90-7.71 (m, 2H, Ar), 7.70-7.45 , J = 16.8 Hz, H- 17), 5.23 (d, 1 H, J = 16.8 Hz, H-17), 2.03-1.81 (m, 2 H, H-19), 1.79-1.08 (br s, 18 H), 1.06-0.92 (m, 12H), 0.80-0.70 (m, 6H). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 171.7, 157.8, 155.5, 155.5, 152.0. 152.0. 151.2, 149.4, 145.0. 132.1, 130.6, 130.0. 128.7, 128.4, 127.9, 119.9, 98.2, 82.7, 81.5, 79.7, 75.2, 65.7, 33.2, 28.3, 27.6, 7.7, 7.2, 6.4.

The second _{^{diastereomer: 1 H NMR (CDCl 3,}} 400 MHz) δ 8.79 (br s, 1 H, Ar), 8.23 (d, 1 H, J = 8.4 Hz, Ar), 8.01 (br d, 1 H, 1H), 7.65-7.76 (m, 2H, Ar), 7.65 (br t, 1H, J = 8.4 Hz, Ar), 7.52 H-5), 5.61 (d, 1H, J = 16.8 Hz, H-17), 5.22 (d, 1H, J = 16.8 Hz, H-17), 2.03-1.82 -19), 1.76-1.08 (br s, 18 H), 1.04-0.92 (m, 12 H), 0.80-0.70 (m, 6 H). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 171.5, 157.9, 155.5, 155.5, 152.3, 152.0. 151.2, 149.4, 145.1, 132.1, 130.6, 130.0. 128.7, 128.4, 127.9, 119.9, 98.2, 82.9, 81.5, 79.6, 75.2, 65.8, 33.3, 28.3, 27.4, 7.8, 7.2, 6.4.

Example III

: 5-di- t -Butoxycarbonylhydrazino-20-OH-camptothecin < / RTI >

It is dissolved in butoxycarbonyl-hydrazino -20 OTES- kaemto camptothecin (0.050 g, 0.072 mmol) first diastereomer is of the anhydrous THF (4 mL) with stirring under an inert environment, - 5-di- t Et ₃ N.3HF (0.088 mL, 0.542 mmol) is then added thereto. The reaction mixture is reacted at room temperature for 35 hours and the disappearance of the reagent (hexane / AcOEt = 3/2) can be monitored by TLC. The solvent is evaporated under vacuum disappears and the residue was purified by flash chromatography (SiO _2, hexane / AcOEt = 3/2), and purification by, and thus the target compound as a light yellow solid (0.041 g, 0.071 mmol, 98 %) .

The product can be further purified by crystallization from CH ₂ Cl ₂ / pentane = 1/50.

¹ H NMR (CDCl ₃ , 400 MHz)? 8.77 (br s, 1 H, Ar), 8.16 (br d, 1 H, J = 8.0 Hz, Ar), 7.97 J = 16.4 Hz, H-17), 5.24 (d, 1H, J = H-19), 1.79-1. 13 (br s, 18 H), 1.03 (t, 3 H , J = 7.6 Hz, Me). ¹³ C NMR (CDCl ₃ , 100 MHz)? 173.7, 157.9, 155.5, 155.5, 152.1, 151.3, 150.7, 149.6, 145.7, 132.3, 130.7, 129.9, 128.7, 127.9, 127.6, 120.0. 97.9, 82.8, 81.6, 79.7, 72.7, 66.1, 31.8, 28.3, 27.7, 7.7.

Example IV

: 5-di- t -Butoxycarbonylhydrazino-20-OH-camptothecin < / RTI >

5-di- t - butoxycarbonyl-hydrazino -20 OTES- kaemto camptothecin second diastereomer (0.050 g, 0.072 mmol) is dissolved in dry of THF (4.5 mL) with stirring under an inert environment, Et ₃ N.3HF (0.088 mL, 0.542 mmol) is then added thereto. The reaction mixture is reacted at room temperature for 35 hours and the disappearance of the reagent (hexane / AcOEt = 3/2) can be monitored by TLC. The solvent is evaporated under vacuum disappears and the residue was purified by flash chromatography (SiO _2, hexane / AcOEt = 3/2), and purification by, and thus the target compound as a light yellow solid (0.040 g, 0.069 mmol, 96 %) .

The product can be further purified by crystallization from CH ₂ Cl ₂ / pentane = 1/50.

¹ H NMR (CDCl ₃ , 400 MHz)? 8.79 (br s, 1 H, Ar), 8.22 (br d, 1 H, J = 8.4 Hz, Ar) J = 16.4 Hz, H-17), 5.26 (d, 1H, J = H-19), 1.79-1.13 (br s, 18 H), 1.03 (t, 3 H, , J = 7.2 Hz, Me). ¹³ C NMR (CDCl ₃ , 100 MHz) δ 173.6, 157.9, 155.4, 155.4, 152.1, 151.3, 150.8, 149.5, 145.6, 132.3, 130.8, 129.8, 128.7, 127.9, 127.8, 119.8, 98.0. 83.0. 81.5, 79.7, 72.7, 66.3, 31.8, 28.3, 27.7, 7.8.

Example V

: Preparation of 5-dibenzyloxycarbonylhydrazino-20-OTES-camptothecin

The camptothecin 20-OTES (0.100 g, 0.216 mmol) was dissolved in anhydrous THF (6 mL) with stirring under an inert atmosphere and then cooled to -78 ° C and 1.0 M LiHMDS in THF (0.281 mL, 0.281 mmol) The solution is put into it. After 20 minutes, dibenzyl azodicarboxylate (0.097 g, 0.324 mmol) in anhydrous THF (2 mL) is added. After 3 h at -78 [deg.] C, the temperature rises to 25 [deg.] C and disappearance of the reagent is monitored by TLC (hexane / AcOEt = 3/1).

The formation of the two diastereomers is observed. After 90 minutes at room temperature, the reaction is cooled by the addition of saturated NH ₄ Cl. The liquid phase is extracted using CH ₂ Cl ₂ (3 x 15 mL) and the organic phases are combined, dried over Na ₂ SO ₄ , filtered under vacuum and condensed. The residue is purified by flash chromatography (SiO _2, hexane / AcOEt = 4/1 or 7/2), and thus to obtain a pale yellow solid (0.161 g, 0.212 mmol, 98 %). The two isomers are separated by further chromatography.

Sequence of elution:

1 ^{st _{diastereomer: 1 H NMR (CDCl 3,}} 400 MHz) δ 8.70 (br s, 1 H, Ar), 8.39 (br s 1 H, Ar), 8.22 (br d, 1 H, J = 7.6 Hz , Ar), 7.95 (br d , 1 H, J = 7.6 Hz, Ar), 7.83 (br t, 1 H, J = 7.6 Hz, Ar), 7.65 (br t, 1 H, J = 7.6 Hz, Ar ), 7.64-7.00 (m, 11 H, Ar + H-14), 6.49 (br s, 1H, H-5), 5.57 (d, 1H, J = 16.4 Hz, H- 4.44 (m, 5 H), 1.98-1.82 (m, 2 H, H-19), 1.02-0.89 (m, 12 H), 0.80-0.70 (m, 6 H). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 171.6, 158.0. 156.3, 156.3, 153.0. 152.2, 151.0. 149.6, 144.8, 135.3, 132.1, 130.6, 130.0. 128.6-127.8 (11 C), 119.9, 98.4, 79.5, 75.2, 68.4, 67.9, 65.6, 33.0. 7.9, 7.2, 6.4.

2 ^nd diastereomer: ¹ H NMR (CDCl ₃ , 400 MHz) δ 8.85 (br s, 1 H, Ar), 8.58 (br s 1 H, Ar), 8.20 (br s, Ar), 7.81 (br t, 1H, J = 7.6 Hz, Ar), 7.63 (br t, 1H, J = 7.6 Hz, Ar), 7.56-6.90 H-14), 6.52 (br s, 1H, H-5), 5.55 (d, 1H, J = 16.8 Hz, H-17), 5.44-4.71

1.98-1.80 (m, 2H, H-19), 1.05-0.90 (m, 12H), 0.81-0.70 (m, 6H). ¹³ C NMR (CDCl ₃ , 100 MHz)? 171.5, 157.9, 156.4, 156.4, 152.9, 152.4, 150.9, 149.4, 144.8, 135.3, 132.1, 130.6, 129.9, 128.6-127.8 , 75.2, 68.4, 67.8, 65.6, 32.9, 7.8, 7.2, 6.4.

Example VI

: The first diastereomer of 5-dibenzyloxycarbonylhydrazino-20-OH-camptothecin

The first diastereomer of 5-dibenzyloxycarbonylhydrazino-20-OTES-camptothecin (0.140 g, 0.184 mmol) was dissolved in anhydrous THF (6 mL) with stirring under inert atmosphere, followed by Et ₃ N.3HF (0.225 mL, 1.380 mmol) is added thereto. The reaction mixture is reacted at room temperature for 52 hours and the disappearance of the reagent (hexane / AcOEt = 1/3) can be monitored by TLC. The solvent is evaporated under vacuum disappears and the residue was purified by flash chromatography and purified by (SiO _2, hexane / AcOEt = 1/1 or 2/3), and thus the target compound as a light yellow solid (0.113 g, 0.175 mmol , 95%). The product can be further purified by crystallization from CH ₂ Cl ₂ / pentane = 1/50.

¹ H NMR (CDCl ₃ , 400 MHz)? 8.67 (br s, 1 H, Ar), 8.39 (br s 1 H, Ar), 8.12 (br d, 1H, J = 7.6 Hz, (br s, 1 H, H), 7.74 (br t, 1H, J = 7.6 Hz, Ar), 7.65-6.66 -5), 5.55 (d, 1H,

J = 16.0 Hz, H-17 ), 5.42-4.44 (m, 5 H), 3.86 (br s, 1 H, OH), 1.92-1.72 (m, 2 H, H-19), 0.95 (t, 3 H, J = 7.6 Hz, Me). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 173.5, 158.0. 156.2, 156.0. 153.0. 150.9, 150.9, 149.5, 145.3, 135.4, 132.2, 130.7, 129.8, 128.7-127.8 (11 C), 119.9, 98.2, 79.6, 72.7, 68.5, 68.0. 65.9, 31.6, 7.8.

Example VII

: The second diastereomer of the crude 5-dibenzyloxycarbonylhydrazino-20-OH-camptothecin

The second diastereomer of 5-dibenzyloxycarbonylhydrazino-20-OTES-camptothecin (0.140 g, 0.184 mmol) was dissolved in anhydrous THF (6 mL) with stirring under inert atmosphere, followed by Et ₃ N.3HF (0.150 mL, 0.921 mmol) is added thereto. The reaction mixture is reacted at room temperature for 55 hours and the disappearance of the reagent (hexane / AcOEt = 3/2) can be monitored by TLC. The solvent is evaporated under vacuum disappears and the residue was purified by flash chromatography (SiO _2, hexane / AcOEt = 1/1), and purification by, and thus the target compound as a light yellow solid (0.113 g, 0.175 mmol, 95 %) . The product can be further purified by crystallization from CH ₂ Cl ₂ / pentane = 1/50.

¹ H NMR (CDCl ₃ , 400 MHz)? 8.71 (br s, 1 H, Ar), 8.34 (br s 1 H, Ar), 8.18 , Ar), 7.79 (br t, 1H, J = 7.6 Hz, Ar),

H-5), 5.53 (d, 1H, J = 16.4 Hz, H-17), 5.44-4.48 (br s, m, 5H), 3.87 (br s, 1H, OH), 1.90-1.70 (m, 2H, H-19), 0.99 (t, 3H, J = 7.6 Hz, Me). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 173.4, 158.0. 156.3, 156.1, 153.0. 151.0. 150.9, 149.6, 145.3, 135.5, 132.3, 130.8, 129.8, 128.7-127.8 (11 C), 119.8, 98.4, 79.5, 72.7, 68.5, 67.8, 66.0. 31.6, 7.7.

Example VIII

: Preparation of 4,5-dihydro-triazole [5,4-c] 16a-deoxo camptothecin TFA salt

5-di- t - butoxycarbonyl-hydrazino -20 OTES- kaemto camptothecin (0.225 g, 0.324 mmol, 1 : 1 diastereomeric mixture) is 1, 2-dichloroethane anhydride with stirring under an inert environment ( DCE) (8 mL), followed by trifluoroacetic acid (TFA) (0.895 mL, 11.67 mmol). The reaction mixture is reacted at room temperature for 20 hours and the disappearance of the reagent (hexane / AcOEt = 1/3) is monitored by TLC and reluxed for 4 hours. The solvent is evaporated under vacuum disappears and the residue was purified by flash chromatography _{(SiO 2, CH 2 Cl 2} / MeOH = 30/1) , and purified by, according to the acetate (trifluoroacetate) trifluoroacetate The desired compound (0.084 g, 0.178 mmol, 55%). The first of the two diastereomers: 1 mixture can be purified further by flash chromatography _{(SiO 2, Toluene / AcOEt =} 1/1).

¹ H NMR (CDCl ₃ , 400 MHz) δ 10.61 (br s, 0.5 H, N 5 -NH═C 16 a), 10.39 (br s, 0.5 H, N 5 -NH═C 16 a) H-7), 8.22-8.15 (m, 1H, Ar), 7.96-7.92 (m, 1H, Ar), 7.88-7.78 Ar),

(M, 2H, Ar, H-17 + N5H), 5.72-5.62 (m, 4.08-3.86 (br s, 1H, OH), 1.96-1.74 (m, 2H, H-19)

1.05-0.98 (t, 3 H, J = 7.6 Hz, Me). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 173.8

(0.5 C), 173.4 (0.5 C), 159.1, 159.0. _{156.7 (q CF 3 COOH),} 156.5 (q CF 3 COOH) 151.5, 151.3, 150.7, 150.5, 150.1, 149.9, 144.8, 144.7, 134.0. 128.4, 128.4, 128.4, 128.2, 128.2, 127.1, 126.9, 120.5, 120.3, 99.1 (2C), 78.9, 78.6, 72.7, 72.7, 66.0 31.7 (2 C), 7.7, 7.7.

Example IX

: Preparation of triazole [5,4-c] 16a-deoxo camptothecin

The 4,5-dihydro-triazole [5,4-c] 16a-deoxo camptothecin TFA salt (0.020 g, 0.042 mmol) was stirred in anhydrous CH ₂ Cl ₂ (4 mL) And then 2,3-dichloro-5,6-dicano-p-benzoquinone (DDQ) (0.025 mg, 0.110 mmol) is added thereto. The reaction mixture is reacted at room temperature for 31 hours and the disappearance of the reagent (CH ₂ Cl ₂ / MeOH = 30/1) can be monitored by TLC. The reaction is cooled by the addition of H ₂ O. The liquid phase is extracted using CH ₂ Cl ₂ (3 x 15 mL) and the organic phases are combined, dried over Na ₂ SO ₄ , filtered under vacuum and condensed. The residue is purified by flash chromatography (SiO ₂ , CH ₂ Cl ₂ / MeOH = 45/1) to give a pale yellow solid (0.014 g, 0.039 mmol, 94%).

_{^{1 H NMR (CDCl 3, 400}} MHz) δ 8.89 (s, 1 H, Ar, H-7), 8.20 (d, 1 H, J = 8.4 Hz, Ar), 8.00 (d, 1 H, J = 8.4 Hz, Ar), 7.88 (t , 1 H, J = 8.4 Hz, Ar), 7.79 (s, 1 H, Ar H-14), 7.69 (t, 1 H, J = 8.4 Hz, Ar), 5.70 ( 1 H, J = 17.2 Hz, H-17), 5.28 (d, 1H, J = 17.2 Hz, H-17), 3.83 H, H-19), 1.08 (t, 3 H, J = 7.6 Hz, Me). _{^{13 C NMR (CDCl 3, 100}} MHz) δ 172.6, 157.4, 152.5, 150.8, 148.9, 143.7, 134.9, 132.5, 132.4, 130.0. 129.5, 128.7, 127.5, 122.6, 121.4, 101.2, 72.4, 66.0. 31.6, 7.7.

Example X: Cell growth inhibition assay

H460 cells from human large-cell lung tumors were cultured in RPMI-1640 medium containing 10% fetal bovine serum. Cell susceptibility was determined by cell growth inhibition assay after exposure to the drug for 1 or 72 hours. Logically growing cells are collected and replicated on 6 well-plates and seeded. 24 hours after the seeding, the cells are exposed to the drug after exposure to the drug and is counted by a Coulter counter (Coulter conter) for the determination of the IC ₅₀ after 72 hours. The IC ₅₀ is defined as the condensation that inhibits cell growth by 50% when compared to untreated control growth.

Example XI: Localisation enzyme-I-dependent DNA rupture analysis

DNA files were obtained from a purified gel of 751-bp BamHI-EcoRI DNA SV40 (Beretta GL, Binaschi M, Zagni AND, Capuani L, Capranico G. "Tethering a type IB topoisomerase to a DNA site by enzyme fusion to a heterologous site- DNA-binding protein domain "(IB) Cancer Res 1999; 59: 3689-97). The DNA fragments were only labeled 3 '. The DNA rupture reaction (20,000 cpm per sample) was performed by adding 10 mM Tris-HCl (pH 7.6), 150 mM KCl, 5 mM MgCl ₂ , 15 μg / mL BSA, 0.1 mM thio Is carried out in 20 ml of dithiothreitol and human genetic recombinant enzyme (full length top 1). The reaction is blocked using 0.5% SDS and 0.3 mg / mL K proteinase for 45 min at 42 < 0 > C. DNA damage persistence is tested at different times by incubation with 10 μM of drug for 30 min followed by addition of 0.6 M NaCl. After precipitation, the DNA was extracted in denaturation buffer (80% formamide, 10 mM NaOH, 0.01 M EDTA, and 1 mg / mL < RTI ID = 0.0 > Dye). ≪ / RTI > All DNA rupture levels were measured by PhosphoImager Model 425 (Molecular Dynamics) (Dallavalle S, Ferrari A, Biasotti B, et al Novel 7-oximinomethyl camptothecin derivatives with potent in vitro and in vivo antitumor activity. J Med Chem 2001; 44: 3264-74).

Persistence of DNA damage (%)

Compounds Time (minutes) 0 One 5 10 Topotecan 100 65 20 10 Camptothecin 100 58 23 20 SN38 100 60 33 28 IDN 6132 100 45 32 20

Claims (8)

In the compounds of formula (I)

(I) R is alkyl, aminoalkyl, hydroxyalkyl, nitrile, alkoximino, aryloximino, silylalkyl; R1 is hydrogen, hydroxy, alkoxy, aminoalkyl; R2 is hydrogen, hydroxy, alkoxy, aminoalkyl, protected hydroxyl; Wherein said alkyl, alkoxy, aminoalkyl or alkoximino groups comprise from 1 to 8 carbon atoms in the form of a straight or branched chain and wherein said aryloxy group has from 5 to 10 carbon atoms, ), Or pharmaceutically acceptable salts, isomers, enantiomers, diastereomers or corresponding mixtures thereof for said compound. The compound of claim 1, wherein the compound of formula (I) a) 4,5-Dihydro-triazole [5,4-c] 16a-deoxo camptothecin and b) Triazole [5,4-c] 16a-deoxo camptothecin Lt; RTI ID = 0.0 > (I) < / RTI > In the process for preparing the compounds of formula (I) according to claim 1, The process comprises steps (a) through (e) substantially as shown in the following table,

here, a) protection of the precursor hydroxy groups; b) 5-position derivatization with N, N-diprotected hydrazine; c) selective conversion of a pyridone ring to a thiopyridone ring; d) elimination of the protecting groups by concomitant cyclization; e) selective aromatization of the pyrazole ring; And wherein R, R1 and R2 have the meanings as defined above and PG is a hydroxy-protecting group. The method of claim 3, Wherein the order of steps (b) and (c) is reversed. A solid tumor or pharmaceutical composition for the treatment of leukemia, comprising a compound of general formula (I) according to claim 1, together with pharmaceutically acceptable carriers and additives. 6. The pharmaceutical composition according to claim 5, A solid tumor or pharmaceutical composition for the treatment of leukemia having the form of a medicament suitable for oral or parenteral administration. Use of the compound of claim 1 or 2 or the pharmaceutical composition of claim 5 or 6 for the preparation of a medicament for the treatment of tumors. 8. The method of claim 7, Methods used for the treatment of tumors such as solid tumors or lungs, ovaries, chest, stomach, liver, prostate, soft tissue sarcoma, head and neck, esophagus, pancreas, colon, rectum, glioblastoma, chronic or acute myelogenous leukemia .